Mallory Denk Bodies (MDBs) are hepatocyte cytoplasmic inclusions that serve as histological hallmark in patients with steatohepatitis (alcoholic and nonalcoholic), but can also be found in other liver disorders including hepatocellular carcinoma and copper metabolism disorders. Recent evidence suggests that the presence of MDBs associates with accelerated liver disease progression. The major constituents of MDBs are the cytoskeletal intermediate filament (IF) proteins, keratin polypeptides 8 and 18 (K8/K18), which form obligate non-covalent K8 and K18 heterodimers that coalesce as tetramers and higher order oligomers. MDB formation is induced by feeding mice with porphyrinogenic drugs, and requires the preferential overexpression of K8 (to establish a K8>K18 state) and K8 crosslinking by transglutaminase. Spontaneous MDBs have been described in mice with a point mutation in the ferrochelatase gene (fch/fch mice), which develop erythropoietic protoporphyria (EPP) due to protoporphyrin deposition in the liver. Also, male mice in one tested strain are more predisposed to MDB accumulation as compared with female mice. The central hypothesis that will be tested in this proposal is that porphyrin deposition, chronic alcohol exposure and genetic gender-associated differences create an environment that promotes MDB formation, and that MDB formation contributes to the hepatocyte ultimate cell fate. The topic covered by this proposal is a novel and understudied area given that the biological significance of MDB formation, and whether MDBs are beneficial or detrimental to hepatocytes, is still not clear despite being first described nearly 100 years ago by Dr. Frank Mallory. Our hypothesis will be tested utilizing three specific aims: (i) Characterize the genetic susceptibility and mechanism of MDB formation in a mouse model of EPP that carries a ferrochelatase gene mutation, (ii) Develop a mouse MDB alcohol model, (iii) Characterize the cell fate of MDB-containing hepatocytes in mouse and human livers. The proposed study combines molecular and biochemical approaches, and state-of-the-art technologies and includes mouse MDB models, as well as human liver MDB molecular analysis. Fundamental findings that we hope to establish include the generation of an alcohol mouse MDB model that is hitherto lacking, the identification of potential gender association with MDB formation in alcoholic liver disease and EPP models, and determining the consequences of MDB formation in individual hepatocytes. Findings from the proposed studies will help clarify whether the pharmacological manipulation of MDBs might be helpful in the treatment of liver diseases. We anticipate that our findings will also contribute to the understanding of other inclusion-associated extra-hepatic disorders, such as myopathies and neuropathies that harbor cytoplasmic IF- containing inclusions. PUBLIC HEALTH RELEVANCE: Patients with a variety of cirrhosis-causing liver diseases develop insoluble deposits in liver cells, termed Mallory-Denk Bodies (MDBs) that clinicians use to help them establish a liver disease diagnosis. The significance of MDBs is poorly understood, but agents that promote their resolution or formation may provide a therapeutic benefit. My proposal will develop mouse models to study MDB formation, and should help us understand in human and mouse livers whether MDBs are protective or detrimental to liver cells.